Optimisation of cell and ex vivo culture conditions to study vascular calcification
Fig 5
Influence of medium composition on ex vivo calcification in aortic rings culture.
A) Influence of the phosphate donor (basic, neutral or acidic). Calcium incorporated into the tunica media of aortic rings assessed by von Kossa staining was higher with neutral phosphate donor buffer (NaH2PO4 / Na2HPO4), P = 0.08. B) Effect of calcium and phosphate concentrations on calcification. A marked increase in calcium content was observed in high phosphate/high calcium conditions, whilst no increase in von Kossa staining positive surface area was observed when compared with high phosphate, suggesting that a significant proportion of calcium was not incorporated into the tunica media (* P<0.01 versus others; ** P<0.001 versus others). C) Effect of alkaline phosphatase on calcification. In 15% FCS supplemented cultures, calcium deposition decreased in the presence of alkaline phosphatase (# P<0.001 for the effect of FCS compared to the same ALK condition; * P<0.01 for the effect of ALK compared to the same FCS condition). (original magnifications x40).